Thank you so much@Mehrbod_Estaki. It is very useful.
And now I go into demultiplexed sequences to be filtered and trimmed, de-noiised, merged. But when I apply dada2 plugin I face a problem. Here is my sequence quality score. and my code and error:
qiime dada2 denoise-paired \
--i-demultiplexed-seqs demultiplexed-seqs.qza
--p-trunc-len-f 290
--p-trunc-len-r 260
--p-trim-left-f 10
--p-trim-left-r 10
--o-representative-sequences rep-seqs-dada2.qza
--o-table table-dada2.qza
--p-n-threads 0
--o-denoising-stats stats-dada2.qza
--verbose
Running external command line application(s). This may print messages to stdout and/or stderr.
The command(s) being run are below. These commands cannot be manually re-run as they will depend on temporary files that no longer exist.
Command: run_dada_paired.R /tmp/tmp7padczzf/forward /tmp/tmp7padczzf/reverse /tmp/tmp7padczzf/output.tsv.biom /tmp/tmp7padczzf/track.tsv /tmp/tmp7padczzf/filt_f /tmp/tmp7padczzf/filt_r 290 260 10 10 2.0 2 consensus 1.0 0 1000000
R version 3.4.1 (2017-06-30)
Loading required package: Rcpp
DADA2 R package version: 1.6.0
- Filtering ............
- Learning Error Rates
2a) Forward Reads
Initializing error rates to maximum possible estimate.
Traceback (most recent call last):
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/q2_dada2/_denoise.py", line 231, in denoise_paired
run_commands([cmd])
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/q2_dada2/_denoise.py", line 36, in run_commands
subprocess.run(cmd, check=True)
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/subprocess.py", line 418, in run
output=stdout, stderr=stderr)
subprocess.CalledProcessError: Command '['run_dada_paired.R', '/tmp/tmp7padczzf/forward', '/tmp/tmp7padczzf/reverse', '/tmp/tmp7padczzf/output.tsv.biom', '/tmp/tmp7padczzf/track.tsv', '/tmp/tmp7padczzf/filt_f', '/tmp/tmp7padczzf/filt_r', '290', '260', '10', '10', '2.0', '2', 'consensus', '1.0', '0', '1000000']' died with <Signals.SIGKILL: 9>.
During handling of the above exception, another exception occurred:
Traceback (most recent call last):
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/q2cli/commands.py", line 274, in call
results = action(**arguments)
File "</home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/decorator.py:decorator-gen-442>", line 2, in denoise_paired
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/qiime2/sdk/action.py", line 231, in bound_callable
output_types, provenance)
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/qiime2/sdk/action.py", line 365, in callable_executor
output_views = self._callable(**view_args)
File "/home/qiime2/miniconda/envs/qiime2-2019.1/lib/python3.6/site-packages/q2_dada2/_denoise.py", line 246, in denoise_paired
" and stderr to learn more." % e.returncode)
Exception: An error was encountered while running DADA2 in R (return code -9), please inspect stdout and stderr to learn more.
Plugin error from dada2:
An error was encountered while running DADA2 in R (return code -9), please inspect stdout and stderr to learn more.
See above for debug info.
so I guess my parameter to trim and trunc is wrong?